In recent years, studying the impact of environmental pollution and toxic molecules on human health has become a research priority. This almost continuous exposure begins during fetal life due to the presence of pollutants in the amniotic fluid (AF) and could be a cause of premature birth. Occurring in 3% of pregnancies, prematurity is a public health problem because of its repercussions on development and learning. Among these pollutants, the phthalates used for many years in most plastics to improve their flexibility are found in high concentrations. In this context, the aim of this work is to develop, optimize and validate a rapid, sensitive and specific analytical method for determining and quantifying six primary phthalate metabolites (MEHP, MiNP, mBzP, MEP, MiBP, and MnBP) and nine primary metabolites of alternative plasticizers (MEHTP, MEHA, MiNCH, 1,2-DEHTM, 1,4-DEHTM, 2,4-DEHTM, 1-MEHTM, 2-MEHTM, and 4-MEHTM) in human AF samples.

Samples were extracted by dispersive liquid-liquid microextraction (DLLME) and UPLC separations were performed on an Acquity UPLC I-Class system (Waters) with a Kinetex Phenyl-Hexyl 150 x 2.1 mm, 1.7µm column (Phenomenex). Each AF sample will first be treated with β-glucuronidase to ensure deglucuronidation of all metabolites. Detection is carried out in Multiple Reaction Monitoring (MRM) mode on a Xevo TQ S-micro mass spectrometer (Waters).

This method can detect traces of plasticizer metabolites with limits of quantification ranged from 0.010 to 0.05 ng/ml. Special care was taken to address two major problems in plasticizer analysis: contamination and chromatographic separation of interfering analogue structures.

This validated quantification method will enable us to analyze 115 AF samples as part of a project funded by the Fondation pour la Recherche Médicale (FRM) studying the link between environmental pollutants (phthalates and alternative plasticizers) and premature rupture of fetal membranes